CN1737158A - The preparation method of creatine kinase bio-sensing device and reagent thereof - Google Patents
The preparation method of creatine kinase bio-sensing device and reagent thereof Download PDFInfo
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Abstract
本发明肌酸激酶生物传感器及其试剂的制备方法，涉及生物传感器技术领域，可以快速检测总肌酸激酶的浓度。本发明肌酸激酶生物传感器，在毛细作用沟道的两电极表面有一纳米材料层，纳米材料层上表面固定有反应试剂；纳米材料层，为铂黑粒子或高分子纳米材料中的一种；以获取高活性的电极表面和高比表面积的纳米材料层。其试剂的制备方法，是先配置两种溶液，再与电子接受体溶液混合而成。当被分析物——含有肌酸激酶的样品——通过毛细作用滴加到试条反应区上后，用电化学生物传感器检测系统可以快速检测出总肌酸激酶的浓度。
The preparation method of the creatine kinase biosensor and the reagent thereof of the present invention relates to the technical field of biosensors, and can rapidly detect the concentration of total creatine kinase. The creatine kinase biosensor of the present invention has a nanomaterial layer on the surface of the two electrodes of the capillary action channel, and a reaction reagent is fixed on the upper surface of the nanomaterial layer; the nanomaterial layer is one of platinum black particles or polymer nanomaterials; In order to obtain a highly active electrode surface and a nanomaterial layer with a high specific surface area. The preparation method of the reagent is that two kinds of solutions are first prepared, and then mixed with the electron acceptor solution. When the analyte—the sample containing creatine kinase—drops onto the reaction zone of the test strip through capillary action, the concentration of total creatine kinase can be quickly detected by the electrochemical biosensor detection system.
Description
Technical Field
The invention relates to the technical field of biosensors, in particular to a test strip for rapidly detecting total creatine kinase by an amperometric biosensor and a manufacturing method of a used reagent.
Technical Field
Creatine Kinase (Creatine Kinase) is present in the cytosol and mitochondria of systemic muscle and brain tissues, and is an important Kinase that has a direct relationship with intracellular energy metabolism, muscle contraction, and regeneration of ATP. Reversibly catalyzing the following reaction:
note: CK: creatine kinase, creatinephoshate: creatine phosphate, ADP: adenosine diphosphate, Creatine: creatine, ATP: adenosine triphosphate, Mg: magnesium ions necessary for creatine kinase reaction activity.
The measurement of creatine kinase activity is one of the basic items for routine examinations in clinical diagnosis of muscle diseases, nerve diseases, encephalopathy, and heart diseases. Cardiovascular diseases, especially myocardial infarction, are a major cause of death in cardiovascular disease. The patients have acute morbidity and high death risk, and the creatine kinase is increased within a few hours after myocardial infarction, so that the creatine kinase of the patients is necessary to be rapidly diagnosed. In addition, the creatine kinase level of prostate cancer, lung cancer and neuroblastoma patients is higher than that of normal people. While hyperglycemia in diabetic patients inhibits the expression of kinase activity in muscle tissue,decreased creatine kinase activity is a consequence of the development of diabetes or a cause of further development of the diabetic condition. And the creatinase kinase detection is one of the important indexes of the physical training intensity of athletes. Under normal conditions, the muscle cell membrane has complete structure and normal function, and Creatine Kinase (CK) rarely penetrates out of the cell membrane. Training and the micro-damage caused by training can increase the creatine kinase activity in the serum. Serum creatine kinase is an important sensitive index for evaluating muscle stimulation and skeletal muscle fine injury and adaptation and recovery thereof.
Currently, the commonly used methods for measuring serum creatine kinase are enzyme-coupled kit method, dry biochemical analysis method and immunoassay method, and the methods all need expensive matched detection instruments. The instrument itself has no design function for emergency treatment. The detection instrument is easily interfered by other substances in serum due to the adoption of the principle of an optical colorimetry and the like, has errors in detection, and cannot detect whole blood.
The invention content is as follows:
the invention aims to provide a portable biosensor creatine kinase test strip which is rapid, accurate and convenient, can be matched with a portable small instrument and can be used for whole blood detection, aiming at the application of outgoing training detection and the like of patients with myocardial infarction acute diagnosis and athletes.
To achieve the above object, the present invention provides a creatine kinase biosensor, comprising the following structure:
a pair of parallel electrodes and electrode leads are arranged on the upper surface of the insulating substrate;
the two electrodes are respectively a working electrode and a counter electrode, and an insulating layer is covered above the two electrodes;
the insulation layer is transversely provided with a groove;
the packaging layer is covered above the groove;
the space enclosed by the insulating substrate, the groove and the packaging layer forms a capillary action channel, two ends of the capillary action channel are provided with capillary sample inlets, the capillary action channel is an electrode reaction area, the surfaces of two electrodes of the capillary action channel are provided with a nanometer material layer, and the upper surface of the nanometer material layer is fixed with a reaction reagent layer; the nano material layer is one of platinum black particles or high polymer nano materials; so as to obtain the electrode surface with high activity and the nanometer material layer with high specific surface area.
The creatine kinase biosensor comprises an electron acceptor, a coupling reaction combined reagent, a buffer solution, an enzyme activator, an enzyme reducer, an anticoagulant and a surfactant.
The creatine kinase biosensor is characterized in that the high-molecular nano material is high-molecular sodium carboxymethyl cellulose, wherein the content of the high-molecular carboxymethyl cellulose is 0.1-1%.
The creatine kinase biosensor, the coupled reaction combined reagent and the analyte react to generate current corresponding to the concentration of the total creatine kinase of the analyte;
the electron acceptor is any one of the group consisting of ferricyanate, methylene blue, ferrocene and derivatives thereof, para-benzoquinone, phenazine methyl sulfate, indophenol and derivatives thereof, and β -naphthoquinone-4-potassium sulfonate;
the buffer solution is any one of the group consisting of phosphate buffer solution, TRIS buffer solution and MES buffer solution;
the enzyme activator is magnesium acetate or magnesium chloride;
the enzyme reducing agent is one of N-acetylcysteine, reduced glutathione or a substance containing sulfhydryl;
the anticoagulant is any one of heparin, oxalate, citrate and EDTA;
the surfactant is any one of TritonX-100, diethanolamide, alkylolamide phosphate, alcohol ether carboxylate, monoalkyl phosphate, sodium dodecyl sulfate or alkylphenol polyoxyethylene.
The creatine kinase biosensor, wherein the coupling reaction combined reagent comprises: creatine phosphate, glucose, Adenosine Monophosphate (AMP), Adenosine Diphosphate (ADP), coenzyme II (NADP) or coenzyme II (NAD), hexokinase or glucokinase, glucose-6-phosphate dehydrogenase, diaphorase.
The creatine kinase biosensor has the following concentration ranges of an electron acceptor: 10 mM-100 mM, buffer solution concentration range: 10-100 mM, enzyme activator concentration range: 0.1-10 mM, concentration range of enzyme reducing agent: 5-20 mM, anticoagulant concentration range: 0.5-5 mM, surfactant concentration range: 0.1 to 0.5 percent.
A preparation method of a reagent for a creatine kinase biosensor comprises the following steps: a) firstly, mixing creatine phosphate and glucose to prepare a solution R1; b) preparing a solution R2 by mixing all or part of Adenosine Monophosphate (AMP), Adenosine Diphosphate (ADP), coenzyme II (NADP) or coenzyme II (NAD), hexokinase or glucokinase, glucose-6-phosphate dehydrogenase and diaphorase, a buffer solution, an enzyme activator, an enzyme reducer, an anticoagulant and a surfactant; c) after thesurface of the electrode is activated, uniformly mixing the solutions R1 and R2 with the electron acceptor solution, and then uniformly dripping the mixture on the electrode nano material layer; d) drying the semi-finished product obtained in the step c), and then covering the semi-finished product with an insulating material to form a packaging layer; e) the array of strips is cut into individual strips by a slitter and then vacuum packaged for use.
The preparation method of the reagent is characterized in that the electrode surface is activated in the step c), and the electrode surface is rapidly activated by using a plasma method.
And d) drying the semi-finished product obtained in the step c) in a drying oven at the temperature of 35-37 ℃ for 20-25 minutes, and covering the packaging layer with an insulating material.
The creatine kinase biosensor has a detection system adopting an amperometric method.
Compared with the current products at home and abroad, the product of the invention has many advantages. The test strip and the introduction method are compatible with a series of commercialized portable current-mode instruments, so that the test strip can be carried about and is simple and quick to detect; because the production process of the disposable test strip adopts an advanced Micro Electro Mechanical System (MEMS) process, the reliability and consistency of detection are ensured, and the production cost of the test strip is reduced; the simple and effective enzyme immobilization method is adopted, the principle of electrochemical current detection is adopted, and compared with the principle of a colorimetric method, the principle is accurate, so that the production of the test strip is simpler and more convenient; the test strip adopts a capillary design, so that the sample introduction is faster, the blood consumption is less (3 mu l), the response of the system is fast, and the sensitivity is high.
Drawings
FIG. 1 is a schematic diagram of a two-electrode system designed according to the present invention in layers;
fig. 2 is a plan view of an electrode of the present invention.
FIG. 3 is a comparison graph of cyclic voltammetry of carboxymethyl cellulose (CMC) with different concentrations in the examples of the present invention.
Fig. 4 is a graph of creatine kinase real-time response for an embodiment of the present invention.
FIG. 5 is a graph of creatine kinase linearity for an embodiment of the present invention.
The specific implementation mode is as follows:
the method for preparing the sensor strip electrode of the invention refers to the applied patent: 00410030214.X and 200410031948. X.
The technique of rapidly activating the electrode surface by plasma method is described in the patent application 02154524.3.
The invention provides a method for assembling a layer of polymer nano-particles in a surface reaction zone of an electrode. Hydrophilic macromolecular sodium carboxymethyl cellulose with different concentrations is uniformly dripped on the surface of the electrode, and after the electrode is dried at the temperature of 37 ℃, electrode nano layers with different apertures are obtained. The method further enhances the hydrophilicity of the electrode surface and increases the effective surface area of the electrode. Not only is beneficial to the immobilization of the enzyme reagent, but also increases the electrochemical activity ofthe electrode.
The present invention provides a method for immobilizing a reactive agent on a nanoparticle.
The invention fixes the reaction reagent on the electrode to form a reagent layer, the reaction reagent fixed on the macromolecule nano layer comprises an electron acceptor, a combined reagent which reacts with the analyte and can generate coupling reaction of current corresponding to the concentration of the analyte, and also comprises buffer solution, enzyme activator, reducing agent, anticoagulant and surfactant.
The reaction principle on which the reagent combination of the invention is based is as follows:
wherein the Creatine phosphate Creatine; creatinne Creatine; CK creatine kinase; glucose; glucose-6-phosphate; gluconolactone-6-phosphate 6-phosphoglucolactone; HK hexokinase; GPD phosphoglucose dehydrogenase; NADPH NADP+(coenzyme II) reduced/oxidized nicotinamide adenine dinucleotide phosphate; DIA diaphorase; fe (CN)6 4-/Fe(CN)6 4-Reduced/oxidized potassium ferricyanide
Firstly, mixing creatine phosphate and glucose with proper concentration to prepare solution R1, mixing Adenosine Monophosphate (AMP), Adenosine Diphosphate (ADP), coenzyme II (NADP), hexokinase, glucose-6-phosphate dehydrogenase, diaphorase, magnesium acetate,reduced glutathione, EDTA and TritonX-100 with proper concentration to prepare solution R2, after the surface of an electrode is activated, uniformly mixing the solutions R1 and R2 and a potassium ferricyanate solution, and then uniformly dripping the solution on an electrode polymer nano layer by using a spotting instrument. And drying in a drying oven at 35-37 ℃ for 20-25 minutes, covering and packaging with an insulating material, and cutting the test strip array into single test strips by using a strip cutting machine. And then vacuum packaging is carried out. And (5) standby.
The applicable concentration range of the creatine phosphate in the reagent is 10-1000 mM, and the optimized reaction range is as follows: 50mM to 100 mM. The applicable concentration range of the glucose is 10-1000 mM, and the optimized reaction range is as follows: 50mM to 100 mM. The applicable concentration range of adenosine monophosphate is 1-100 mM, and the optimized reaction range is as follows: 5mM to 20 mM. The application concentration range of the adenosine diphosphate is 0.5-100 mM, and the optimized reaction range is as follows: 1mM to 20 mM. The applicable concentration range of the coenzyme II is 0.5-100 mM, and the optimized reaction range is as follows: 1mM to 20 mM. The applicable concentration ranges of hexokinase, glucose-6-phosphate dehydrogenase and diaphorase are 1-2000U/ml, and the optimized reaction range is as follows: 1-40U/ml.
The electron acceptor with the redox effect is selected from one of the group consisting of ferricyanate, methylene blue, ferrocene and derivatives thereof, para-benzoquinone, phenazine methyl sulfate, indophenol and derivatives thereof and β -naphthoquinone-4-potassium sulfonate, electrons generated in the enzyme reaction process are transferred to the surface of an electrode from an enzyme reaction center through an electron mediator, so that the response speed and the detection sensitivityof the current enzyme sensor are improved, the reaction voltage is reduced, the working potential is reduced to 0.2V due to the fact that the electron acceptor is the ferricyanate, the interference of other active substances in blood is reduced, and the application concentration range of the ferricyanate is 0.1-300 mM, and the optimized reaction range is 10-100 mM.
The buffer solution is one selected from the group consisting of phosphate buffer solution, TRIS buffer solution and MES buffer solution; the buffer solution is phosphate buffer solution. The buffer solution is used for providing a reaction environment with stable pH value, the pH value of the optimal pH value of the reaction is 6-8, and the concentration range is as follows: 10 to 100 mM.
In the creatine kinase reaction, magnesium ions are necessary, and the enzyme activator is magnesium acetate or magnesium chloride. Selecting magnesium acetate, wherein the applicable concentration range is as follows: 1-100 mM, and the optimal concentration range is as follows: 5 to 20 mM.
Most creatine kinase in serum exists in an inactive form, and must be activated for accurate quantitative determination of the activity, and a commonly used enzyme reducing agent is one of sulfhydryl-containing substances such as N-acetylcysteine, reduced glutathione and the like; the enzyme reducing agent is reduced glutathione. Because the sulfhydryl compound has poor stability in solution, the reduced glutathione needs to be freshly prepared solution. The applicable concentration range is as follows: 0.1-10 mM, and the optimal concentration range is as follows: 0.1 to 5 mM.
In the whole blood detection process, blood is easy to coagulate, and generally the blood starts to coagulate within 2-5 minutes, so that an anticoagulant needs to be added to a reagent in order to make the reaction smoothly proceed. The anticoagulant is one selected from heparin, oxalate, citrate and EDTA; the anticoagulant is EDTA. The applicable concentration range is as follows: 0.1-10 mM, and the optimal concentration range is as follows: 0.5 to 5 mM.
The surfactant is TritonX-100. 0.05 to 1% of a nonionic surfactant is added. Preferably 0.1-0.5% surfactant.
The various objects, methods, features and advantages of the present invention as described above will be more fully understood from the following description taken in conjunction with the accompanying drawings and examples.
Example 1
Fig. 1 and 2 are two electrode diagrams designed by the invention. Firstly, thin film gold electrodes 2 and 3 are prepared on an insulating base material 1 by adopting a Micro Electro Mechanical System (MEMS) process, a second insulating layer 4 is adhered on the thin film gold electrodes 2 and 3, a capillary channel 7 is arranged in the second insulating layer 4, and the capillary channel 7 forms a reaction area. (5 is a packaging layer, 6 is an electrode lead, 10 is a capillary sample inlet). Then, after the electrodes 2 and 3 are activated by an ion cleaning machine, a layer of polymer carboxymethyl cellulose (CMC) nano-particle layer 8 is assembled in the reaction area of the electrodes 2 and 3, namely the capillary action channel 7, and is dried in an electrothermal blowing dry box at 37 ℃ for 20 minutes and taken out for standby.
Then reagents were added, the reagents being: creatine phosphate: 100 mM; glucose: 50 mM; magnesium acetate: 20 mM; adenosine Monophosphate (AMP): 20 mM; adenosine Diphosphate (ADP): 5 mM; nicotinic Adenosine Dinucleotide Phosphate (NADP): 5 mM; glucose 6-phosphate dehydrogenase: 4U/ml; hexokinase: 4U/ml; myocardial xanthase: 2U/ml; glutathione: 3 mM; EDTA:1 mM; TritonX-100: 0.2 percent; potassium ferricyanate: 25 mM; phosphate buffer: 0.01M.
Mu.l of the above reagent solution was dropped into the capillary channel 7, coated with a layer of polymeric carboxymethyl cellulose (CMC) nanoparticle layer 8, and dried in a 37 ℃ dry box for 20 minutes to form a reagent layer 9. The strip is removed from the dry box and capped with a transparent insulating encapsulant 5 to form capillary channels 7 and capillary sample inlets 10 at both ends. The array of strips is cut into individual strips with a slitter. And then vacuum packaging is carried out.
Example 2
After the prepared electrodes 2 and 3 were washed with ionized water, 0.5% and 1% of polymeric carboxymethyl cellulose (CMC) were added dropwise and dried in an electrothermal blowing dry box at 37 ℃ for 20 minutes. After the sample was taken out, the voltammetry curve was measured. In fig. 3, the electrodes 2 and 3 are assembled with different concentrations of carboxymethyl cellulose (CMC): (1)0 percent; (2)0.5 percent; (3) cyclic voltammogram at 1%. Scanning rate: 100mV/s, solution base phase: 0.01M phosphate buffer(pH7.4)，0.03MK3Fe(CN)6. As can be seen from fig. 3, as the concentration of the polymeric carboxymethyl cellulose (CMC) increases, the current thereof increases, but as the concentration of the polymeric carboxymethyl cellulose (CMC) increases, the polymeric nano-film thickens, the electron transfer rate decreases, and the reaction speed is affected; the background reaction current increases. Therefore, it is necessary to select an appropriate concentration. Generally, the concentration is 0.1-1%, which is determined according to specific conditions. The surface area of the electrodes 2 and 3 assembled with the polymer nano-film is obviously increased, and a liquid reaction test is addedAfter the reagent is added, the nanometer material layer 8 and the reagent form a sol layer, so that larger reaction current is generated, and the reaction sensitivity is increased.
Example 3
A voltage of +0.2V is applied between the working and reference electrodes of the strip. The mixed reagent is dripped on the electrodes 2 and 3 modified with the high molecular material sodium carboxymethyl cellulose (CMC) nano layer 8, after the reaction current (background current) is stable, creatine kinase buffer solutions with different activities are added to react with the reagent and the high molecular nano layer 8 to generate current, the current flows out through an electrode lead 6, and the current corresponding to the activity of total creatine kinase can be quickly detected on an electrochemical biosensor detection system. The current produced is proportional to the activity of the kinase. And exhibits a good linear dependence over a wide range. Therefore, the sensor test strip has good application value. See fig. 4a, which is a real-time response curve of a biosensor to different concentrations of creatine kinase. Creatine kinase activity: (1) 800U/ml; (2) 400U/ml; (3) 80U/ml; (4) 8U/ml; (5) 800U/L; (6) 400U/L; (7) 80U/L; (8) 40U/L; (9) 8U/L. Fig. 4b is an enlarged view of the dotted line portion of fig. 4 a. Fig. 5 is a creatine kinase linearity chart. Fig. 5a shows the high activity range of creatine kinase, reaction time: 140 seconds; fig. 5b shows creatine kinase base activity range, reaction time: 280 seconds.
The present invention has been described in terms of the preferred embodiment, which is intended in an illustrative rather than in a limiting sense. Many modifications and variations of the present invention are possible in light of the above teachings. Thus, within the scope of the appended claims, the invention may have other implementations than those described above. For example: other combined reagent formulas, a reagent layer covered with a filter membrane to eliminate interference of red blood cells for detection of whole blood creatine kinase, an electrode reaction area which can adopt optical detection method, other nano-particles and non-nano material modification, a three-electrode form, a preparation form of a thick-film electrode and the like.
Claims (11)
1. A creatine kinase biosensor comprising the structure:
a pair of parallel electrodes and electrode leads are arranged on the upper surface of the insulating substrate;
the two electrodes are respectively a working electrode and a counter electrode, and an insulating layer is covered above the two electrodes;
the insulation layer is transversely provided with a groove;
the packaging layer is covered above the groove;
insulating substrate, slot and the space that the encapsulated layer encloses constitute the capillary channel, and capillary channel both ends are the capillary introduction port, and this capillary channel is electrode reaction district, its characterized in that:
a nanometer material layer is arranged on the surfaces of the two electrodes of the capillary action channel, and a reaction reagent layer is fixed on the upper surface of the nanometer material layer; the nano material layer is one of platinum black particles or high polymer nano materials; so as to obtain the electrode surface with high activity and the nanometer material layer with high specific surface area.
2. The creatine kinase biosensor of claim 1, wherein: the reaction reagent comprises an electron acceptor, a coupling reaction combined reagent, a buffer solution, an enzyme activator, an enzyme reducer, an anticoagulantand a surfactant.
3. The creatine kinase biosensor of claim 1, wherein: the polymer nano material is polymer sodium carboxymethyl cellulose, wherein the content of the polymer sodium carboxymethyl cellulose is 0.1-1%.
4. The creatine kinase biosensor of claim 2, wherein: the coupling reaction combined reagent reacts with the analyte and can generate current corresponding to the concentration of the total creatine kinase of the analyte;
the electron acceptor is any one of the group consisting of ferricyanate, methylene blue, ferrocene and derivatives thereof, para-benzoquinone, phenazine methyl sulfate, indophenol and derivatives thereof, and β -naphthoquinone-4-potassium sulfonate;
the buffer solution is any one of the group consisting of phosphate buffer solution, TRIS buffer solution and MES buffer solution;
the enzyme activator is magnesium acetate or magnesium chloride;
the enzyme reducing agent is one of N-acetylcysteine, reduced glutathione or a substance containing sulfhydryl;
the anticoagulant is any one of heparin, oxalate, citrate and EDTA;
the surfactant is any one of TritonX-100, diethanolamide, alkylolamide phosphate, alcohol ether carboxylate, monoalkyl phosphate, sodium dodecyl sulfate or alkylphenol polyoxyethylene.
5. The creatine kinase biosensor of claim 4, wherein: the coupling reaction combined reagent comprises: creatine phosphate, glucose, Adenosine Monophosphate (AMP), Adenosine Diphosphate (ADP), coenzyme II (NADP) or coenzyme II (NAD), hexokinase or glucokinase, glucose-6-phosphate dehydrogenase, diaphorase.
6. The creatine kinase biosensor of claim 4, wherein: concentration range of the electron acceptor: 10 mM-100 mM, buffer solution concentration range: 10-100 mM, enzyme activator concentration range: 0.1-10 mM, concentration range of enzyme reducing agent: 5-20 mM, anticoagulant concentration range: 0.5-5 mM, surfactant concentration range: 0.1 to 0.5 percent.
7. The creatine kinase biosensor of claim 5, wherein: in the coupling reaction combined reagent, the concentration range of the creatine phosphate is 10-1000 mM; the concentration range of the glucose is 10-1000 mM; the concentration range of the adenosine monophosphate is 1-100 mM; the concentration range of the adenosine diphosphate is 0.5-100 mM; the concentration range of the coenzyme II is 0.5-100 mM; the concentration ranges of hexokinase, glucose-6-phosphate dehydrogenase and diaphorase are 1-2000U/ml.
8. A preparation method of a reagent used by a creatine kinase biosensor is characterized by comprising the following steps: comprises the following steps: a) firstly, mixing creatine phosphate and glucose to prepare a solution R1; b) preparing a solution R2 by mixing all or part of Adenosine Monophosphate (AMP), Adenosine Diphosphate (ADP), coenzyme II (NADP) or coenzyme II (NAD), hexokinase or glucokinase, glucose-6-phosphate dehydrogenase and diaphorase, a buffer solution, an enzyme activator, an enzyme reducer, an anticoagulant and a surfactant; c) after the surface of the electrode is activated, uniformly mixing the solutions R1 and R2 with the electron acceptor solution, and then uniformly dripping the mixture on the electrode nano material layer; d) drying the semi-finished product obtained in the step c), and then covering the semi-finished product with an insulating material to form a packaging layer; e) the array ofstrips is cut into individual strips by a slitter and then vacuum packaged for use.
9. The method of claim 7, wherein: and c) activating the electrode surface in the step c), namely rapidly activating the electrode surface by using a plasma method.
10. The method of claim 7, wherein: and d), drying the semi-finished product obtained in the step c) in a drying oven at the temperature of 35-37 ℃ for 20-25 minutes, and covering the packaging layer with an insulating material.
11. The creatine kinase biosensor of claim 1, wherein: the detection system adopts a current method for detection.
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| CN1916623B (en) * | 2005-08-17 | 2013-05-29 | 希森美康株式会社 | Reagent for creatine kinase activity test |
| CN104485232A (en) * | 2014-12-18 | 2015-04-01 | 中国科学院上海硅酸盐研究所 | Preparing method of counter electrode for dye-sensitized solar cell |
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| JP2775055B2 (en) * | 1989-02-08 | 1998-07-09 | 新日本無線株式会社 | Biosensor |
| GB9421468D0 (en) * | 1994-10-18 | 1994-12-07 | Surface Active Ltd | Biosensor |
| ATE315225T1 (en) * | 1996-03-29 | 2006-02-15 | Byk Gulden Italia Spa | AUTOMATIC DIAGNOSTIC DEVICE |
| CN100389322C (en) * | 2002-11-27 | 2008-05-21 | 中资汉脉（北京）生物技术有限公司 | Nano silicon membrane biological sensor |
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| CN1916623B (en) * | 2005-08-17 | 2013-05-29 | 希森美康株式会社 | Reagent for creatine kinase activity test |
| CN104485232A (en) * | 2014-12-18 | 2015-04-01 | 中国科学院上海硅酸盐研究所 | Preparing method of counter electrode for dye-sensitized solar cell |
| CN104485232B (en) * | 2014-12-18 | 2017-06-06 | 中国科学院上海硅酸盐研究所 | A kind of preparation method of used by dye sensitization solar battery to electrode |
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